Live biological processes have previously only been observed by light microscopy at low spatial resolution (~200nm), while the molecular details of biological samples have only been studied at high-resolution (~1 nm) in the arrested (fixed or frozen/cryo) state by transmission electron microscopy (TEM). Here we propose to merge these two applications, by purchasing, modifying and operating a Dynamic TEM (DTEM) that can "film" live biological processes in real-time with sub-molecular resolution. The DTEM is a recent TEM development, in which the electron source uses a pulsed laser that produces short bursts of electrons in the 5s or ns range via photoemission, permitting "snapshot images" (single pulse) and "movies" (multiple pulses) of dynamic processes to be recorded. Samples will be loaded into the Bio-DTEM using a specially designed liquid stage, where fluid (live) biological samples can be observed. The proposed instrument will also incorporate latest developments in aberration correction and electron phase- shifting technology, to deliver images at unprecedented contrast and clarity. Combined, these technological advancements will allow for the first time to visualize live biological samples in liquid solutions at a resolution allowing the identification of sub-molecular details. This proposal links unique technical expertise at Lawrence Livermore National Laboratory (LLNL) in the design and operation of the DTEM with extensive biomedical research programs at UC-Davis. The aim of this proposal is to reach a new paradigm for biomedical imaging that will be available in a user facility for biomedical researchers. Initially, research will focus on research currently funded by NIH, ranging from an analysis of the chronology and key-players in chromosome mechanics (maintenance and repair), mitosis and mitochondrial fusion/fission events, to the study of membrane pore formation by pentraxins or the detailed processes in membrane protein crystallization. PUBLIC HEALTH RELEVANCE: The proposed Bio-DTEM will allow the study of dynamic events at unprecedented resolution in cellular, molecular and pharmacological research. Biological processes that so far have been studied by confocal light microscopy, will be observable with 100 times improved resolution. This instrument will enable milestone advancements in our understanding of cancer, bacterial or viral infections, and basic biological processes.